(Solution) SCIE1046: Activity 6.2 ELISA

COURSE  

SCIE1046: Fundamental of Microbiology with Lab

. About the Lab

Learning Objectives:

  • Explain the principle of different ELISA techniques.
  • Apply sandwich ELISA to quantify protein samples.
  • Analyze the standard curve of ELISA experiment.
  • Understand the function of reagents and equipment used in ELISA.
  • Describe the basic troubleshooting process of ELISA.

Estimated Length: 50 minutes to 1 hour

 

MAKE THE CONNECTION

The background information in section 2 was adapted from the following Microbiology lecture course Tutorials:
5.4.1 Antibody Production and Laboratory Testing
5.4.5 EIA, FEIA, and ELISA Testing

 

  1. Background Information

The following background information will be helpful as you prepare for the simulation.

In this lab simulation, the term hapten is used to reference the size of molecules. A hapten is a small molecule that binds with a protein carrier to elicit antibody production. Within the lab, four types of ELISA testing are covered. The type of ELISA testing used will vary based on whether you are assessing antibody or antigen activity and whether you are measuring small or large molecules.

There are conflicting definitions on what constitutes a small vs a large molecule. In the context of pharmacology, a small molecule is composed of less than 1,000 atoms, while a larger molecule is composed of more than 1,000 atoms.

 

TERM TO KNOW

This glossary term is important to know and will help you during the Activity.

Hapten

A small molecule that binds with a protein carrier to elicit antibody production.

 

2a. Data and Graphs

Raw data are the actual data values used in a set of scores used for assessment. During this lab simulation, you will be using raw data from the ELISA to create graphs. Graphs are mathematical models that plot data using points positioned above a number line. A graph will visually present data in mass numbers so that we can easily observe where data cluster and how they distribute around a mean, or average.

In a normal distribution, also called a standard distribution or standard curve, data are evenly distributed around the mean. A standard distribution often presents as a bell curve.

During an ELISA, multiple dilutions with a range of concentrations are conducted. This results in lots of raw data that are graphed to create a standard curve.

 

TERMS TO KNOW

These glossary terms are important to know and will help you during the Activity.

Raw Data

The actual data values used in a set of scores used for assessment.

Graphs

Mathematical models that plot data using points positioned above a number line. A graph visually presents data in mass numbers so that we can easily observe where data cluster and how they distribute around a mean, or average.

Normal Distribution

Data are evenly distributed around the mean, often presented as a bell curve. Also called standard distribution or standard curve.

 

2b. Serial Dilution

In order to create the multiple dilutions needed for an ELISA test, you will want to use a serial dilution technique. Serial dilutions are made by creating the same dilution step over and over using a previous dilution as the input for the next dilution to be conducted. Since the dilution fold is the same in each step, the dilutions create a constant ratio between any subsequent dilutions.

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